CytosolCytosol

HepatocyteHepatocyteApoB

DegradationDegradation

5' 3'

ApoB mRNA ApoB mRNA TranslationTranslation

ER

Mem

bran

eE

R M

embr

ane

InsulinIRS1/IRS2

IR

Insu

lin S

igna

ling

Path

way

Insu

lin S

igna

ling

Path

way

Plasma

Plasma

VLDLVLDL

Insulin Signaling & VLDL Overproduction Insulin Signaling & VLDL Overproduction in Insulin Resistancein Insulin Resistance

PI3-Kinase

VLDL AssemblyVLDL Assembly

PIP3/Phosphorylation CascadePIP3/Phosphorylation Cascade

Akt/PKBAkt/PKB

PTP-1BPTP-1B

3'

D

Insu

lin R

ecep

tor p

rote

in M

ass

(sca

nnin

g un

its/ m

g pr

otei

n x1

0-3)

Control Fructose-fed

Control Fructose-fed0

50

100

150

200

250

IRS

-1 P

rote

in M

ass

(sca

nnin

g un

its/m

g pr

otei

n x1

0-3)

Control Fructose-fed

Control Fructose-fed0

40

80

120

E F Control Fructose-fed

0

20

40

60

80

100

120

Control Fructose-fed

IRS

-2 M

ass

(per

cent

of c

ontro

l)

C

Pho

spho

ryla

ted

IRS

-2(r

elat

ive

to b

asal

leve

l of c

ontro

l)

020406080

100120140160180

_ + _+

Insulin

+ + + +Insulin _ _ _ _

Control Fructose-fed

ControlFructose-fed

Impaired Hepatic Tyrosine Phosphorylation of Insulin Receptor, IRS-1, and IRS-2

in Fructose-Fed HamstersA

Pho

spho

ryla

ted

Insu

lin R

ecep

tor M

ass

(sca

nnin

g un

its/m

g pr

otei

n x1

0-3)

Control Fructose-fed

0

100

200

300

400

500

600

_ + _+

Insulin

+ + + +Insulin _ _ _ _

*

B

_ + _+

Insulin

Pho

spho

ryla

ted

IRS

-1(s

cann

ing

units

/mg

prot

ein

x10-3

)

Control Fructose-fed

0

20

40

60

80

100

120

Control

Fructose-fed

+ + + +Insulin _ _ _ _

P= 0.04

P= 0.009

P= 0.02P= 0.03

P= 0.029

*

*

B

0

50

100

150

200

PTP

-1B

Mas

s(p

erce

nt o

f con

trol)

Control Fructose-fed

Control Fructose-fed

C

0

100

200

300

PTP

-1B

Act

ivity

(per

cent

of c

ontro

l)

Control Fructose-fed

A

0

20

40

60

80

100

120

PI 3

-Kin

ase

Act

ivity

(per

cent

of c

ontro

l)

Control Fructose-fed

* *****

D + + + +Insulin

020406080

100120140

Insu

lin-In

duce

d A

kt S

erin

ePh

osph

oryl

atio

n (%

of c

ontr

ol)

Control Fructose-fed

Insulin

_ + _ +

E

Insulin

Insu

lin-In

duce

d A

kt T

hreo

nine

Ph

osph

oryl

atio

n (%

of c

ontr

ol)

Control Fructose-fed

020

40

60

80

100120

_ + _ +

+ + + +

Insulin

F Control Fructose-fed

20406080

100120140160

Akt

Pro

tein

Mas

s (p

erce

nt o

f con

trol)

0Control Fructose-fed

****

***

*

Evidence for Reduced PI-3 Kinase Activity, Reduced Akt Phosphorylation, & Enhanced PTP-1B Mass & Activity

0

20

40

60

80

100

120

0 20 40 60 80

Vanadate (M)

Rec

over

ed C

ellu

lar A

poB

(rel

ativ

e to

unt

reat

ed)

B

0

20

40

60

80

100

120

0 20 40 60 80Vanadate (M)

Rec

over

ed L

abel

ed M

edia

Apo

B(r

elat

ive

to u

ntre

ated

)

C

Vanadate (M)

Rec

over

ed T

otal

Apo

B(r

elat

ive

to u

ntre

ated

)

0

20

40

60

80

100

120

0 20 40 60 80

D

Vanadate (M) 0 10 40 80

IR-pY

0

100

200

300

400

500

600

0 20 40 60 80Vanadate (M)In

sulin

Rec

epto

r Pho

spho

ryla

tion

(rel

ativ

e to

unt

reat

ed)

A

0 10 20 40 80Vanadate (M)

Cellular ApoB

Secreted ApoB

E

Inhibition of Cellular Phosphatase Activity with Vanadate Enhances Insulin Signaling and Reduces ApoB Secretion

IR-IR-ppYY

Cellular ApoBCellular ApoB

ApoB StabilityApoB Stability

ApoB SecretionApoB Secretion